The long-term objectives of the proposed research are to understand the genetic and cellular mechanisms which regulate B lymphocyte activation, differentiation, and expression of antibody diversity. The present research focuses of the activation requirements and antibody responses of two functionally and phenotypically distinct B cell subsets which correspond to different stages in B cell development: the Lyb5- and Lyb5+ B cell subsets. The sepcific aims are (1) to determine whether Iyb5- B cells can be stimulated by certain determinants if they are presented as type 1 rather than type 2 antigens; (2) to determine whether the antibodies produced by Lyb5- and Lyb5+ B cells share idiotypy, and if so, in which isotypes; and (3) to determine whether cross-linking surface Ia molecules and surface Ig molecules on B cells by antibody serves as an inductive signal, and if so, in which B cell subset. The methodology includes the in vitro splenic focus system for B cell cloning; iso-electric focusing and Western blot analysis; production of hybridoma antibodies and anti-idiotypic antibodies; B cell proliferative assays and assays for Ig secretion; and radioimmunoassays for antibody levels, isotypes, and idiotypes.